Protein Structure and Folding
Solution Structures of a Cyanobacterial Metallochaperone: INSIGHT INTO AN ATYPICAL COPPER-BINDING MOTIF*

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The Atx1 copper metallochaperone from Synechocystis PCC 6803, ScAtx1, interacts with two P1-type copper ATPases to supply copper proteins within intracellular compartments, avoiding ATPases for other metals en route. Here we report NMR-derived solution structures for ScAtx1. The monomeric apo form has a βαββα fold with backbone motions largely restricted to loop 1 containing Cys-12 and Cys-15. The tumbling rate of Cu(I)ScAtx1 (0.1–0.8 mm) implies dimers. Experimental restraints are satisfied by symmetrical dimers with Cys-12 or His-61, but not Cys-15, invading the copper site of the opposing subunit. A full sequence of copper ligands from the cell surface to thylakoid compartments is proposed, considering in vitro homodimer liganding to mimic in vivo liganding in ScAtx1-ATPase heterodimers. A monomeric high resolution structure for Cu(I)ScAtx1, with Cys-12, Cys-15, and His-61 as ligands, is calculated without violations despite the rotational correlation time. 2JNH couplings in the imidazole ring of His-61 establish coordination of Nϵ2 to copper. His-61 is analogous to Lys-65 in eukaryotic metallochaperones, stabilizing Cu(I)S2 complexes but by binding Cu(I) rather than compensating charge. Cys-Cys-His ligand sets are an emergent theme in some copper metallochaperones, although not in related Atx1, CopZ, or Hah1. Surface charge (Glu-13) close to the metal-binding site of ScAtx1 is likely to support interaction with complementary surfaces of copper-transporting ATPases (PacS-Arg-11 and CtaA-Lys-14) but to discourage interaction with zinc ATPase ZiaA and so inhibit aberrant formation of copper-ZiaA complexes.

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The atomic coordinates and structure factors (code 1SB6) have been deposited in the Protein Data Bank, Research Collaboratory for Structural Bioinformatics, Rutgers University, New Brunswick, NJ (http://www.rcsb.org/).

*

This work was supported by the European Community Grant SPINE QLG2-CT-2002-00988, (“Structural Proteomics in Europe”) and by MIUR COFIN 2003 “Il Ruolo degli Ioni Metallici nei Processi Metabolici.” The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

The on-line version of this article (available at http://www.jbc.org) contains Tables 1 and 2 and Figs. 1, 2, 3, 4, 5.

Supported by the Biotechnology and Biological Sciences Research Council.